Investigation of KATP Channel Endocytosis and Cell Surface Density by Biotinylation and Western Blotting

2001 ◽

Vol 8 (5) ◽

pp. 932-936 ◽

Cited By ~ 3

Author(s):

Cynthia L. Bristow

Keyword(s):

Human Immunodeficiency Virus ◽

Cell Surface ◽

Chemokine Receptors ◽

Surface Density ◽

Human Leukocyte ◽

Absolute Number ◽

Mononuclear Phagocytes ◽

Leukocyte Elastase ◽

Immunodeficiency Virus

ABSTRACT The absolute number of CD4+ lymphocytes in blood is prognostic for disease progression, yet the cell surface density of CD4 receptors or chemokine receptors on a single cell has not previously been found to be predictive of human immunodeficiency virus (HIV) infectivity outcome. It has recently been shown that human leukocyte elastase (HLE) and its ligand α1 proteinase inhibitor (α1PI; α1 antitrypsin) act as HIV fusion cofactors. The present study shows that decreased HIV infectivity is significantly correlated with decreased cell surface density of HLE but not with decreased CD4 nor chemokine receptors. In vitro HIV infectivity outcome in this study was predicted by the surface density of HLE on mononuclear phagocytes but not on lymphocytes. The set point HLE surface density was in part determined by α1PI. Decreased circulating α1PI was correlated with increased cell surface HLE and with increased HIV infectivity. The correlation of HIV infectivity outcome with surface HLE and circulating α1PI supports the utility of these HIV cofactors in diagnostic analysis and therapeutic intervention.

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CD63 is a component of Weibel-Palade bodies of human endothelial cells

Blood ◽

10.1182/blood.v82.4.1184.bloodjournal8241184 ◽

1993 ◽

Vol 82 (4) ◽

pp. 1184-1191 ◽

Cited By ~ 3

Author(s):

UM Vischer ◽

DD Wagner

Keyword(s):

Endothelial Cells ◽

Cell Surface ◽

Western Blotting ◽

Vascular Endothelial Cells ◽

Secretory Granules ◽

Positive Staining ◽

Cell Fractionation ◽

Glycosylation Pattern ◽

Adhesion Proteins ◽

Shape Distribution

Weibel-Palade bodies are secretory granules of vascular endothelial cells specialized in the storage of von Willebrand factor (vWF) and P- selectin, two adhesion proteins that can be rapidly mobilized to the cell surface by exocytosis in response to thrombin or other agonists. In this study, we attempted to identify additional components of Weibel- Palade bodies by raising monoclonal antibodies to these granules, purified by cell fractionation. One antibody, 2C6, was found to be specific for CD63, a membrane glycoprotein previously described in the lysosomes of platelets and other cell types. The immunopurified 2C6 antigen was recognized by an anti-CD63 reference antibody, 2.28, by Western blotting. Also, the biosynthetic profile of the 2C6 antigen in endothelial cells showed a nascent molecular mass and a glycosylation pattern identical to that of CD63. Immunofluorescence staining with 2C6 showed the lysosomes, and also elongated structures identified as Weibel-Palade bodies by their shape, distribution, and positive staining with anti-vWF antibodies, CD63 was also found by Western blotting of subcellular fractions highly enriched in Weibel-Palade bodies. Our results indicate that CD63 colocalizes with vWF and P- selectin in the Weibel-Palade bodies of endothelial cells, and together with these adhesion proteins it could be rapidly expressed on the cell surface in areas of vascular injury and inflammation.

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A generic cell surface ligand system for studying cell-cell recognition

10.1101/546846 ◽

2019 ◽

Author(s):

Eleanor M Denham ◽

Michael I Barton ◽

Susannah M Black ◽

Marcus J Bridge ◽

Ben de Wet ◽

...

Keyword(s):

Cell Surface ◽

Surface Density ◽

Cell Receptor ◽

Surface Receptors ◽

Ligand System ◽

Ligand Interactions ◽

A Cell ◽

Surface Ligand ◽

Receptor Biology ◽

Cell Cell

AbstractDose-response experiments are a mainstay of receptor biology studies and can reveal valuable insights into receptor function. Such studies of receptors that bind cell surface ligands are currently limited by the difficulty in manipulating the surface density of ligands at a cell-cell interface. Here we describe a generic cell surface ligand system that allows precise manipulation of cell surface ligand densities over several orders of magnitude. We validate the system for a range of immunoreceptors, including the T cell receptor (TCR), and show that this generic ligand stimulates via the TCR at a similar surface density as its native ligand. This system allows the effect of surface density, valency, dimensions, and affinity of the ligand to be manipulated. It can be readily extended to other receptor-cell surface ligand interactions, and will facilitate investigation into the activation of, and signal integration between, cell surface receptors.

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Extracellular K+ concentration controls cell surface density of IKr in rabbit hearts and of the HERG channel in human cell lines

Journal of Clinical Investigation ◽

10.1172/jci39027 ◽

2009 ◽

Vol 119 (9) ◽

pp. 2745-2757 ◽

Cited By ~ 106

Author(s):

Jun Guo ◽

Hamid Massaeli ◽

Jianmin Xu ◽

Zongchao Jia ◽

Jeffrey T. Wigle ◽

...

Keyword(s):

Cell Surface ◽

Cell Lines ◽

Human Cell ◽

Surface Density ◽

Herg Channel ◽

Human Cell Lines ◽

K Concentration

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The relationship between released soluble FceRI-alpha and its cell surface density on human basophils

PLoS ONE ◽

10.1371/journal.pone.0245942 ◽

2021 ◽

Vol 16 (1) ◽

pp. e0245942

Author(s):

Donald MacGlashan

Keyword(s):

Molecular Weight ◽

Flow Cytometry ◽

Cell Surface ◽

Surface Density ◽

Alpha Subunit ◽

Constant Amount ◽

Western Blots ◽

Ige Mediated ◽

The Relationship ◽

Human Basophils

Background The IgE-mediated activation of mast cells and basophils results in the secretion of many substances, including the release of FceRI-alpha subunit. This released alpha subunit can bind IgE and it may act as a down-regulator of subsequent IgE-dependent reactions. However, previous studies do not observe loss of the mass of FceRI-alpha associated with the cells, at least not for human basophils. This study was designed to understand the basis for the discordant observations. Methods Purified human basophils were stimulated with multiple activating secretagogues and supernatants were examined for histamine and released FceRI-alpha. In addition, cell surface IgE densities (occupied and unoccupied) were measured by flow cytometry and total cellular content of mature and immature FceRI-alpha determined with Western blots. Results Released FceRI-alpha, on average, represented 7% of the total surface FceRI before the reaction. The molecular weight of the soluble FceRI-alpha was approximately 54 kD, larger than immature subunit and somewhat smaller than surface subunit. In addition, 1) release ceased long before internalized FceRI-alpha was processed, 2) release was insensitive to Bafilomycin A, 3) release was independent of the starting density of FceRI and 4) release occurred more effectively with non-IgE-dependent stimuli, FMLP or C5a. Conclusions There appears to be relatively constant amount of nearly mature FceRI-alpha that is susceptible to secretion events induced by any form of stimulation. The amount, on average, represents about 7% of the mature form of FceRI-alpha.

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Heat shock protein 70-reactivity is associated with increased cell surface density of CD94/CD56 on primary natural killer cells

Cell Stress and Chaperones ◽

10.1379/1466-1268(2003)008<0348:hspria>2.0.co;2 ◽

2003 ◽

Vol 8 (4) ◽

pp. 348 ◽

Cited By ~ 71

Author(s):

Catharina Gross ◽

Ingo G.H. Schmidt-Wolf ◽

Srinivas Nagaraj ◽

Robert Gastpar ◽

Joachim Ellwart ◽

...

Keyword(s):

Heat Shock ◽

Natural Killer Cells ◽

Cell Surface ◽

Heat Shock Protein ◽

Natural Killer ◽

Heat Shock Protein 70 ◽

Surface Density ◽

Killer Cells

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Protein Antigen in Serotype k Streptococcus mutans Clinical Isolates

Journal of Dental Research ◽

10.1177/154405910808701001 ◽

2008 ◽

Vol 87 (10) ◽

pp. 964-968 ◽

Cited By ~ 21

Author(s):

K. Nakano ◽

R. Nomura ◽

H. Nemoto ◽

J. Lapirattanakul ◽

N. Taniguchi ◽

...

Keyword(s):

Infective Endocarditis ◽

Dental Caries ◽

Cell Surface ◽

Streptococcus Mutans ◽

Western Blotting ◽

Surface Protein ◽

Clinical Isolates ◽

Cell Surface Protein ◽

Western Blotting Analysis ◽

Blotting Analysis

Streptococcus mutans, a major pathogen of dental caries and infective endocarditis, is classified into serotypes c, e, f, and k, with serotype k strains recently reported to be frequently detected in persons with infective endocarditis. Thus, we hypothesized that common properties associated with infective endocarditis are present in those strains. Fifty-six oral S. mutans strains, including 11 serotype k strains, were analyzed. Western blotting analysis revealed expression of the 3 types of glucosyltransferases in all strains, while expression of the approximately 190-kDa cell-surface protein (PA) was absent in 12 strains, among which the prevalence of serotype k (7/12) was significantly high. Furthermore, cellular hydrophobicity and phagocytosis susceptibility were lower in the group of serotype k strains. These results indicate that the absence of PA expression, low cellular hydrophobicity, and phagocytosis susceptibility are common bacterial properties associated with serotype k strains, which may be associated with virulence for infective endocarditis.

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